Production of specific antibodies against SARS-coronavirus nucleocapsid protein without cross reactivity with human coronaviruses 229E and OC43

Abstract

Severe acute respiratory syndrome (SARS) is a life-threatening disease for which accurate diagnosis is essential. Although many tools have been developed for the diagnosis of SARS, false-positive reactions in negative sera may occur because of cross-reactivity with other coronaviruses. We have raised polyclonal and monoclonal antibodies (Abs) using a recombinant form of the SARS virus nucleocapsid protein. Cross-reactivity of these anti-SARS Abs against human coronavirus (HCoV) 229E and HCoV OC43 were determined by Western blotting. The Abs produced reacted with recombinant SARS virus nucleocapsid protein, but not with HCoV 229E or HCoV OC43.

Fig. 1

RT-PCR of cell lysates infected with human coronavirus (HCoV) 229E and HCoV OC43. A: HCoV 229E specific RT-PCR, B: HCoV OC43 specific RT-PCR. The results of RT-PCR were consistent with virally infected MRC-5 cell. Lane M: 100 bp DNA ladder, Lane MRC-5: normal MRC-5 cell lysates, Lane 229E: HCoV 229E infected MRC-5 cell lysates, Lane OC43: HCoV OC43 infected MRC-5 cell lysates.

Fig. 2

Western blotting for detecting cross reactivity of polyclonal antibody (Ab) and monoclonal Ab with HCoVs 229E and OC43. A: SDS-PAGE, B: reacted with polyclonal Ab, C: reacted with monoclonal Ab. Purified recombinant N protein (Lanes N), HCoV 229E infected cell lysates (Lanes 229E) and HCoV OC43 infected cell lysates (Lanes OC43) were run in SDS PAGE 12% gels with molecular weight markers in Lane M.